Evaluation of imipenem for prophylaxis and therapy of Yersinia pestis delivered by aerosol in a mouse model of pneumonic plague.

It has been previously shown that mice subjected to an aerosol exposure to Yersinia pestis and treated with ß-lactam antibiotics after a delay of 42 h died at an accelerated rate compared to controls. It was hypothesized that endotoxin release in antibiotic-treated mice accounted for the accelerated death rate in the mice exposed to aerosol Y. pestis. Imipenem, a ß-lactam antibiotic, binds to penicillin binding protein 2 with the highest affinity and produces rounded cells. The binding of imipenem causes cells to lyse quickly and thereby to release less free endotoxin. Two imipenem regimens producing fractions of time that the concentration of free, unbound drug was above the MIC (fT>MIC) of approximately 25% (6/24 h) and 40% (9.5/24 h) were evaluated. In the postexposure prophylaxis study, the 40% and 25% regimens produced 90% and 40% survivorship, respectively. In the 42-h treatment study, both regimens demonstrated a 40 to 50% survivorship at therapy cessation and some deaths thereafter, resulting in a 30% survivorship. As this was an improvement over the results with other ß-lactams, a comparison of both endotoxin and cytokine levels in mice treated with imipenem and ceftazidime (a ß-lactam previously demonstrated to accelerate death in mice during treatment) was performed and supported the original hypotheses; however, the levels observed in animals treated with ciprofloxacin (included as an unrelated antibiotic that is also bactericidal but should cause little lysis due to a different mode of action) were elevated and significantly (7-fold) higher than those with ceftazidime.

Multiple roles of Myd88 in the immune response to the plague F1-V vaccine and in protection against an aerosol challenge of Yersinia pestis CO92 in mice.

The current candidate vaccine against Yersinia pestis infection consists of two subunit proteins: the capsule protein or F1 protein and the low calcium response V protein or V-antigen. Little is known of the recognition of the vaccine by the host's innate immune system and how it affects the acquired immune response to the vaccine. Thus, we vaccinated Toll-like receptor (Tlr) 2, 4, and 2/4-double deficient, as well as signal adaptor protein Myd88-deficient mice. We found that Tlr4 and Myd88 appeared to be required for an optimal immune response to the F1-V vaccine but not Tlr2 when compared to wild-type mice. However, there was a difference between the requirement for Tlr4 and MyD88 in vaccinated animals. When F1-V vaccinated Tlr4 mutant (lipopolysaccharide tolerant) and Myd88-deficient mice were challenged by aerosol with Y. pestis CO92, all but one Tlr4 mutant mice survived the challenge, but no vaccinated Myd88-deficient mice survived the challenge. Spleens from these latter nonsurviving mice showed that Y. pestis was not cleared from the infected mice. Our results suggest that MyD88 appears to be important for both an optimal immune response to F1-V and in protection against a lethal challenge of Y. pestis CO92 in F1-V vaccinated mice.

Human anti-plague monoclonal antibodies protect mice from Yersinia pestis in a bubonic plague model.

Yersinia pestis is the etiologic agent of plague that has killed more than 200 million people throughout the recorded history of mankind. Antibiotics may provide little immediate relief to patients who have a high bacteremia or to patients infected with an antibiotic resistant strain of plague. Two virulent factors of Y. pestis are the capsid F1 protein and the low-calcium response (Lcr) V-protein or V-antigen that have been proven to be the targets for both active and passive immunization. There are mouse monoclonal antibodies (mAbs) against the F1- and V-antigens that can passively protect mice in a murine model of plague; however, there are no anti-Yersinia pestis monoclonal antibodies available for prophylactic or therapeutic treatment in humans. We identified one anti-F1-specific human mAb (m252) and two anti-V-specific human mAb (m253, m254) by panning a naïve phage-displayed Fab library against the F1- and V-antigens. The Fabs were converted to IgG1s and their binding and protective activities were evaluated. M252 bound weakly to peptides located at the F1 N-terminus where a protective mouse anti-F1 mAb also binds. M253 bound strongly to a V-antigen peptide indicating a linear epitope; m254 did not bind to any peptide from a panel of 53 peptides suggesting that its epitope may be conformational. M252 showed better protection than m253 and m254 against a Y, pestis challenge in a plague mouse model. A synergistic effect was observed when the three antibodies were combined. Incomplete to complete protection was achieved when m252 was given at different times post-challenge. These antibodies can be further studied to determine their potential as therapeutics or prophylactics in Y. pestis infection in humans.

CpG oligodeoxynucleotides augment the murine immune response to the Yersinia pestis F1-V vaccine in bubonic and pneumonic models of plague.

The current U.S. Department of Defense candidate plague vaccine is a fusion between two Yersinia pestis proteins: the F1 capsular protein, and the low calcium response (Lcr) V-protein. We hypothesized that an immunomodulator, such as CpG oligodeoxynucleotide (ODN)s, could augment the immune response to the plague F1-V vaccine in a mouse model for plague. CpG ODNs significantly augmented the antibody response and efficacy of a single dose of the plague vaccine in murine bubonic and pneumonic models of plague. In the latter study, we also found an overall significant augmentation the immune response to the individual subunits of the plague vaccine by CpG ODN 2006. In a long-term, prime-boost study, CpG ODN induced a significant early augmentation of the IgG response to the vaccine. The presence of CpG ODN induced a significant increase in the IgG2a subclass response to the vaccine up to 5 months after the boost. Our studies showed that CpG ODNs significantly augmented the IgG antibody response to the plague vaccine, which increased the probability of survival in murine models of plague (P<0.0001).